AbstractNeural tube is a structure within the developing vertebrate embryo that gives rise to the central nervous system consisting of the brain and the spinal cord. Within the neural tube, there are pools of specific neural progenitors that give rise to specialised types of neurons. Those progenitors are organised in distinct domains and the process of the establishment of such a pattern ventrally is tightly regulated by Shh morphogen, released by cells of organising centres in the ventral midline of the embryo, namely the floor plate and the notochord. In recent years, a lot of research has been carried out to understand how cells interpret the Shh signal and translate this information in order to adopt a specific neuronal fate.
The Notch pathway plays a crucial role during neurogenesis of both invertebrate and vertebrate embryos. It has also been demonstrated that the Notch and Shh signalling pathways can interplay in various developmental contexts. In this project, I have studied the novel role of Notch signalling in the patterning of the ventral neural tube on the dorsal-ventral axis and the interplay between Notch and Shh signalling. Analysis of gene expression patterns in genetically modified mouse embryos revealed that loss of Notch pathway leads to decreased size of ventral domains, namely p3 and floor plate. Moreover, repression of Notch with small inhibitor compound demonstrated in in vitro assays that progenitors of neural explants have lower sensitivity to the Shh signal. Likewise, activation of Notch in the ventral midline of mouse embryo, at the developmental stage when this pathway becomes attenuated in the floor plate, induced expression of p3 marker Nkx2.2 and Shh target gene Ptch1. In addition, the Notch target gene in chicken embryo HAIRY2 was demonstrated to be also the target of Shh signalling in the ventral midline and to play role in induction and maintenance of the p3 fate. Missexpression of this gene in the ventral neural tube promotes p3 fate at the expense of floor plate at motorneuron fates. Further understanding of Notch and Shh interaction was achieved by using cell based assays with mouse and chicken fibroblast cells, NIH3T3 and DF1 respectively, well-established in vitro models for studying Shh signalling. It was demonstrated that changes in Notch signalling modulate the localisation of key components of the Shh pathway, namely Smoothened and Patched1, to primary cilia. Activation of Notch increases ciliary localisation of Smoothened, whereas inhibition of Notch decreases exit of Patched1 from cilia upon Shh stimulation. Finally yet importantly, I find that high Notch activity increases the length of primary cilia in the ventral neural tube of mouse embryos in floor plate and p3 progenitors, and in NIH3T3 cell culture. The findings of this project demonstrate that Notch signalling pathway plays important role in the early patterning of the ventral neural tube and provides new information into understanding of floor plate formation and cellular interpretation of Shh morphogen gradient.
|Date of Award||2015|
|Sponsors||Biotechnology and Biological Sciences Research Council|
|Supervisor||Kim Dale (Supervisor)|